(a) Field
The subject matter disclosed generally relates to gel electrophoresis. More specifically, the subject matter disclosed relates to an electrolyte solution for extending the useful electrophoresis life of an electrophoresis gel during gel electrophoresis containing at least one zwitterion, and water.
(b) Related Prior Art
Gel electrophoresis is a common procedure for the separation of biological molecules, such as deoxyribonucleic acid (DNA), ribonucleic acid (RNA), polypeptides and proteins. In gel electrophoresis, the molecules are separated into bands according to the rate at which an imposed electric field causes them to migrate through a filtering gel.
The basic apparatus used in this technique consists of a gel enclosed in a glass tube, sandwiched as a slab between glass or plastic plates, or poured in a plastic tray. The gel has an open molecular network structure, defining pores which are saturated with an electrically conductive buffered solution. These pores through the gel are large enough to admit passage of the migrating macromolecules.
The gel is placed in a chamber in contact with buffer solutions which make electrical contact between the gel and the cathode or anode of an electrical power supply. A sample containing the macromolecules and a tracking dye is placed on top of the gel. An electric potential is applied to the gel causing the sample macromolecules and tracking dye to migrate toward the bottom of the gel. The electrophoresis is halted just before the tracking dye reaches the end of the gel.
The most common buffer system employed for the separation of proteins is the Laemmli buffer system consists of 0.375 M tris(hydroxy methyl)amino-methane (Tris), titrated to pH 8.8 with HCl, in the separating gel. The stacking gel consists of 0.125 M Tris, titrated to pH 6.8. The anode and cathode running buffers contain 0.024 M Tris, 0.192 M glycine, 0.1% SDS. Many different gel separation materials have been disclosed, with different compositions, pH characteristics, voltage requirements, etc. The goal of most of the recent innovations in the field has been to provide an electrophoresis gel which can be used to perform a faster, more accurate, more stable, or therefore more versatile electrophoresis.
A number of different gel buffer systems have been proposed for use at or around neutral pH that do not involve the use of the Tris-HCl Glycine buffer system of Laemmli.
For example, U.S. Pat. No. 6,096,182 to Updyke et al. discloses an electrophoresis gel at a neutral pH. The advantage of producing such a gel is that the gel system is stable, with reduced reactivity and increased shelf life.
U.S. Pat. No. 5,464,517 to Hjerten et al. discloses an electrophoresis buffer which has a high buffering capacity and low electrical conductivity. The advantage of this type of buffer, particularly in capillary electrophoresis, is that it allows the separation to be performed at a higher voltage and consequently more quickly.
A majority of innovations have focused on improving electrophoresis by proposing new recipes for the gel buffer.
Currently, a major obstacle in the production and sale of pre-cast electrophoresis gels is their rather short shelf life of about 3 months. For example, for pre-cast polyacrylamide gels, it is believed that their degradation is a consequence of the hydrolysis of amide groups to form partially anionic carboxylic acid derivatives under basic conditions. Therefore, the high pH (e.g. pH 8.0 to 9.5) is believed to lower the stability of the gels upon storage. The hydrolysis is believed to lead to loss of resolution of the separated molecules, reduced migration distance of the separated molecule, and the reduced intensity of protein staining.
Therefore, there is a need for reagents that will extend the useful electrophoresis life of gels that have gone pass their normal expiry date (shelf-life).